This research project is aimed at defining the organization of the adenovirus genome in terms of the location and function of the virus genes which control virus development during infection, and at defining the requirements for adeno gene products in oncogenic transformation of rodent cells. Temperature-sensitive (ts) and host-range (hr) mutants of human adenovirus 5 and of Ad2ionND1 have been isolated, and these are being characterized genetically and physiologically. Many of the mutants have been genetically mapped, and ts mutations have been mapped physically by restriction analysis of intertypic recombinants. Recently we have developed improved methods of transfection for adeno DNA and DNA-protein complex, and the resulting enhanced efficiency of transfection has allowed us to refine the physical mapping limits of many ts and hr mutations by a marker rescue technique using restriction fragments of DNA. We aim to use the technique to introduce mutated or modified fragments into the Ad5 genome. The mutants are being characterized for DNA and protein synthesis, T-antigen production, DNA infectivity, etc., in both lytic and non-lytic interactions with cells. The hr mutants are defective for transformation of rat cells and may have mutations in the genes needed for initiation and maintenance of transformation.